Summary Neisseria gonorrhoeae is the etiological agent of the sexually transmitted infection (STI) gonorrhea, a high morbidity disease worldwide with ~ 106 million cases annually. Like many human pathogens this organism must adapt to environments encountered during infection, including low pH and varying oxygen and iron levels. Tight control of gene expression in the gonococcus is mediated in part by the Fur protein, which binds to specific DNA sequences leading to either activation or repression of a repertoire of genes. We recently reported that the gonococcal Fur regulon extends to additional regulatory proteins, which together contribute to gonococcal mechanisms of pathogenesis. We also established that subsets of Fur regulated genes are expressed during natural gonococcal infection in women relative to during growth in vitro. Based collectively on our results we hypothesize that the gonococcal Fur regulon extends to additional regulatory networks that are expressed and regulated in the female genital tract. The studies proposed in this application will define how Fur-mediated regulation is extended through control of additional regulators that are crucial to natural infection. Our analysis will focus on regulatory proteins and sRNAs that are 1) Controlled via Fur (directly or indirectly) and 2) Regulated during natural infection in humans. We term these ?gonococcal regulators expressed in vivo? (GREIV). The following Aims are proposed: Aim 1. To define the repertoire of Fur-controlled gonococcal regulatory proteins and sRNAs expressed during natural infection of the female genital tract. Aim 2. To characterize the role of newly identified Fur-controlled GREIV in N. gonorrhoeae interactions with epithelial cells and innate immune cells. Aim 3. To fully define gonococcal global regulatory networks expressed during natural infection of the female genital.